cloning, expression and library construction for hiv-1 tat protein

background: designing novel therapeutic agents has been a critical challenge for hiv disease. materials and methods: in current study a dna sequence which was encoded the tat protein was synthesized and inserted in pet 28 vector. vector was cloned in bl21-de3 e. coli and cultured in tb media. after protein expression, recombinant tat protein was purified by nta affinity chromatography. the tat purified protein efficiency and confirmed by sds-page and western blot, respectively. we were immunized the camel against hiv-1 tat recombinant protein to made a camelid antibody library. total rna was extracted from camel lymphocytes and vhh fragments synthesized and amplified using rt-pcr and nested- pcr methods by special primers. results: the 350- 450 bp vhh gene fragment was produced by rt-pcr and nested- pcr and extracted from agarose gel 1%. then gel extraction was performed and pure fragments were inserted in hen-4 vector by t4 dna ligase. conclusion: the library can be applied for biopanning and isolation of nanobody against hiv-1 tat protein. nanobody small size may be a useful drug for treatment of hiv disease because give them the potency of the recognizing the cryptic epitopes of tat and neutralized the virus.